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Toxicity to aquatic algae and cyanobacteria

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Reference
Endpoint:
toxicity to aquatic algae and cyanobacteria
Type of information:
experimental study
Adequacy of study:
key study
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: Study meets criteria for Klimisch score 1 because it follows an internationally recognised mthod and was carried out to Good Laboratory Practice.
Qualifier:
according to
Guideline:
EPA OTS 797.1050 (Algal Toxicity, Tiers I and II)
Deviations:
yes
Remarks:
The determination of algistatic vs algicidal effects was omitted as the study was completed as a limit test
GLP compliance:
yes
Analytical monitoring:
yes
Details on sampling:
Total organic carbon (TOC) measurements of initial (0 hours) high (1000 mg/L), low (100 mg/L) and control test solutions and at test termination (96 hours), conducted on each replicaate control and treatment vessel.
Vehicle:
no
Details on test solutions:
The 100, 300 and 1000 mg/L nominal loadings were prepared using sterile enriched alga growth media. Individual water accommodation fractions were prepared by stirring the vessel for 24 hours with a Teflon coated magnetic stir bar and removing the water phase for the toxicity test.
Test organisms (species):
Pseudokirchneriella subcapitata (previous names: Raphidocelis subcapitata, Selenastrum capricornutum)
Details on test organisms:
TEST ORGANISM
- Common name: Green algae
- Source (laboratory, culture collection): Cells taken from a log-growth phase in-house culture originally purchased from University of Texas at Austin alga collection
Test type:
static
Water media type:
freshwater
Limit test:
yes
Total exposure duration:
96 h
Remarks on exposure duration:
readings taken at 72 hrs also
Post exposure observation period:
Not reported
Hardness:
Not reported
Test temperature:
23.3 to 24°C
pH:
7.5 to 9.8 at 0 hours and 8.3 to 10.1 after 96 hours
Dissolved oxygen:
Not reported
Salinity:
Not relevant
Nominal and measured concentrations:
Control, 100, 300 and 1000 mg/L WAF.

TOC levels were 2-8 mg/L in the control, 2-4 mg/L at 100 mg/L WAF, 2-3 mg/L at 300 mg/L WAF and 2-3 mg/L at 1000 mg/L WAF loading. TOC levels were not considered to be indicative of actual test material concentrations and results are therefore based on nominal loading rates.
Details on test conditions:
TEST SYSTEM
- Test vessel: 250 mL Erlenmeyer flasks containing 100 mL stoppered with foam plugs to reduce the entry of dust randomly placed on an orbital shaker to approximately 100 cycles per minute
- Renewal rate of test solution: Solution was not renewed
- Initial cells density: 10,000 cells/mL
- No. of vessels per concentration (replicates): Three

GROWTH MEDIUM: Sterile enriched alga media adjusted to pH 7.5. Measured TOC and total suspended solids in fresh untreated alga media were <1.0 and <10 mg/L, respectively.

OTHER TEST CONDITIONS
- Sterile test conditions: Yes
- Adjustment of pH: Adjusted to pH 7.5
- Photoperiod: Constant light (24 hours per day)
- Light intensity: 47-50 µEin/m²second

EFFECT PARAMETERS MEASURED:
- Determination of cell concentrations: Visual determination by means of direct microscope examination with a hemocytometer

TEST CONCENTRATIONS
- Test concentrations: Control, 100, 300 and 1000 mg/L WAF
Reference substance (positive control):
no
Key result
Duration:
72 h
Dose descriptor:
NOEC
Effect conc.:
1 000 mg/L
Nominal / measured:
nominal
Conc. based on:
other: WAF loading rates
Basis for effect:
growth rate
Key result
Duration:
72 h
Dose descriptor:
EC50
Effect conc.:
> 1 000 mg/L
Nominal / measured:
nominal
Conc. based on:
other: WAF loading rates
Basis for effect:
growth rate
Key result
Duration:
72 h
Dose descriptor:
EC50
Effect conc.:
> 1 000 mg/L
Nominal / measured:
nominal
Conc. based on:
other: WAF loading rates
Basis for effect:
biomass
Key result
Duration:
96 h
Dose descriptor:
NOEC
Effect conc.:
1 000 mg/L
Nominal / measured:
nominal
Conc. based on:
other: WAF loading rates
Basis for effect:
growth rate
Key result
Duration:
96 h
Dose descriptor:
EC50
Effect conc.:
> 1 000 mg/L
Nominal / measured:
nominal
Conc. based on:
other: WAF loading rates
Basis for effect:
growth rate
Key result
Duration:
96 h
Dose descriptor:
EC50
Effect conc.:
> 1 000 mg/L
Nominal / measured:
nominal
Conc. based on:
other: WAF loading rates
Basis for effect:
biomass
Details on results:
The control response was satisfactory.

At 72-hours, biomass measurements were 80%, 62% and 70% of the control at 100, 300 and 1000 mg/L WAF, respectively. At 96-hours, biomass measurements were 70%, 66% and 88% of the control at 100, 300 and 1000 mg/L WAF, respectively.
Reported statistics and error estimates:
A parametric one-way analysis of variance (ANOVA) and Dunnett's test were used to calculate the no-observed effect level.

The test vessels containing 1,000mg/L WAFs were slightly cloudy at the start of the test and at 24hours. The algal population grew well during the test, resulting in an average of 1,711,000cells/ml in the control (doubling time= 13.0hours).

Cell growth data from the acute toxicity test with the WAF of 3 concentrations of substance and the freshwater alga,Selenastrum capricornutum are provided below with % growth rates compared to control are shown below at time points 0, 24, 48, 72 and 96 hours.

0 (control)

 

1

10

18

123

826

1,555

 

 

2

10

21

145

1,008

1,866

 

 

Mean

10

20

134

917

1,711

 

%

Control

-—

100

 

1

10

15

118

680

1,250

 

 

2

10

18

133

790

1,145

 

 

Mean

10

17

126

735

1,198

 

%

Control

100

85

94

80

70

300

 

1

10

<10

120

506

1,125

 

 

2

10

<10

107

638

1,120

 

 

Mean

10

<10

114

572

1,122

 

%

Control

100

<50

85

62

66

1000

 

1

10

<10

132

654

1,610

 

 

2

10

<10

125

632

1,385

 

 

Mean

10

<10

129

643

1,498

 

%

Control

100

<50

96

70

88

               

                                        

 

Based on the number of cells/ml or the average specific growth rate, the 72 and 96 hour EC50s are >1,000 mg/L.

No effects (size differences, unusual cell shapes, colors, flocculations, adherence of cells to test containers, or aggregation of cells) were noted. Based on the number of cells/ml or the average specific growth rate, the 72 and 96 hour NOEC values are 1,000 mg/L.

Validity criteria fulfilled:
yes
Conclusions:
The test substance was not toxic to freshwater algae at loading rates up to and including 1000 mg/L nominal WAF. The 72 and 96 hour ErC50 and EbC50 are considered to be >1000mg/l
Executive summary:

In a 96-hour acute toxicity study, the cultures of Selenastrum capricornutum (now Pseudokirchneriella subcapitata) were exposed to test substance (115733-09-0) at nominal concentrations of 100, 300 and 1000 mg/l as WAF loadings under static conditions in accordance with the EPA OTS 797.1050. 

 

Daily cell counts were made visually by means of direct microscopic examination with a hemocytometer.

 

This study is classified as reliable without restriction (Klimisch Code 1) and satisfies the guideline requirements for a toxicity study to aquatic algae and cyanobacteria.

  

Results Synopsis

 

Test Organism: Selenastrum capricornutum (now Pseudokirchneriella subcapitata)

Test Type: Static

 

At 72-hours biomass measurements in the treatments were 80%, 62% and 70% of the control at 100, 300 and 1000 mg/L, respectively.

At 96-hours biomass measurements in the treatments were 70%, 66% and 88% of the control at 100, 300 and 1000 mg/L, respectively.

 

72-hour and 96-hour Effective Loading Rate (EL50) = >1000 mg/L nominal WAF.

72-hour and 96-hour No Observed Effect Levels = 1000 mg/L nominal WAF based on both growth rate and biomass.

Description of key information

The toxicity of the substance has been measured experimentally with freshwater algae.  Due to the extremely low water solubility all studies have been conducted using Water Accomodated Fractions and completed as limit tests.  

Key value for chemical safety assessment

EC50 for freshwater algae:
1 000 mg/L
EC10 or NOEC for freshwater algae:
1 000 mg/L

Additional information

The EC50 in freshwater algae is considered to be >1000mg/l. Two of the studies available support this conclusion, while the one remaining study did not exceed 100 mg/l, although the data support the conclusion that the EC50 is above this dose level. As this dose level has been included in the other studies conducted, it is considered acceptable to consider this as supporting data but to conclude that the highest dose level attained from studies considered reliable is both appropriate and adequately supported by the data.

The substance is, therefore, considered not toxic to algae. Nevertheless, adequate controls will be in place during production and use to ensure that the substance is not released to the environment.

The toxicity of the substance has not been assessed using marine algal species, but it is considered likely to be similar to freshwater species.